荷斯坦奶牛杀菌/通透性增加蛋白N端cDNA在大肠杆菌中的融合表达  

Expression of Bactericidal/Permeability-increasing Protein N-terminal cDNA of Hostein Cow in Escherichia coli

作　　者：高恒[1] 彭开松[1] 涂健[1] 祁克宗[1] 江龙海[1] 

机构地区：[1]安徽农业大学动物科技学院,合肥230036

出　　处：《农业生物技术学报》2009年第4期599-602,共4页Journal of Agricultural Biotechnology

基　　金：国家高技术研究与发展计划(863)项目(No.2006AA10Z320);国家自然科学基金项目(No.30871851;No.30800811);安徽省教育厅自然科学研究项目(No.KJ2007A021)资助

摘　　要：参照GenBank中安哥斯牛杀菌/通透性增加蛋白(BPI)序列,设计合成1对引物。应用RT-PCR技术,从荷斯坦奶牛嗜中性粒细胞mRNA中扩增出杀菌/通透性增加蛋白N端,将该序列与原核表达载体pGEX-4T-1连接,构建重组质粒pGEX-4T-1-BPI。重组质粒转化大肠杆菌(Escherichia coli)BL21(DE3),用IPTG诱导表达。结果表明,获得了BPIN端长度为714bp的cDNA序列,序列分析证实在第503位出现了点突变。重组质粒诱导表达后,经SDS-PAGE电泳检测重组蛋白约在52kD处出现特异性蛋白条带。According to the Bactericidal/permeability-increasing Protein （BPI） sequence of Angus calf in GenBank, a pair of primers were designed. The N-terminal cDNA was amplified by RT-PCR from mRNA which was extracted from the polymorphonuclear neutrophils of Hostein cow. The purified cDNA was connective with expression vector pGEX-4T-1 to construct recombinant plasmid pGEX-4T-1-BPI, which was transformed into Escherichia coli BL21. The results showed that a 714 bp fragment was acquired, and there was a basic mutated group in 503rd compared with the reports. The E.coli with recombinant plasmid was induced with IPTG. The products were analyzed by SDS-PAGE, and a new protein of 52 kD was detected. Its molecular weight was the same as expected, showing the purified recombinant BPI was obtained successfully.

关 键 词：杀菌/通透性增加蛋白 表达 荷斯坦奶牛 

分 类 号：S188[农业科学—农业基础科学]